Isolation of a conditional suppressor of leucine auxotrophy in Saccharomyces cerevisiae

Abstract

OR 97403, USAPhenotypically and genotypically (/eu2-3,112) Leu-cells of Saccharomyces cerevisiae gave rise to small colonies on medium devoid of leucine. This only occurred on plates with a high density of Leu-cells or on medium supplemented with limiting quantities of leucine. Cells from these small colonies retained a growth advantage over their parent on limiting leucine supplements even after growth in a non-selecting (rich) medium. Therefore, the growth variants had acquired a heritable change. The phenotype was recessive and due to a change in a nuclear gene unlinked to the LEU2 locus. The phenotype provided a growth advantage only during leucine starvation; growth of the variants was indistinguishable from their parent on medium lacking the other essential supplements (histidine and uracil) required for the growth of the strain.[14C]Leucine uptake assays demonstrated that the variants were better able than their parents to accumulate leucine from their environments, and this ability extended to other hydrophobic amino acids. These results suggest that in the variants an amino acid uptake system has been derepressed rather than there having been reversion or extragenic suppression of the mutation in leucine biosynthesis. We designate the mutant gene responsible for the phenotype /up1 (for leucine uptake). The transport characteristics of the /up1 mutants suggested that LUPl is a regulatory component of an ammonium-regulated hydrophobic amino acid uptake system.

The coevolution of toxin and antitoxin genes drives the dynamics of bacterial addiction complexes and intragenomic conflict

“…In a nitrogen-limiting environment in which amino acids may be prevalent, an amino acid transporter is adaptive. However, in the same environment with toxic amino acid analogues, expression of the transporter may be deleterious or lethal [45]. Similarly, transporters can suppress the effects of mutations in biosynthetic pathways, being adaptive when the essential metabolite cannot be made, but deleterious when a toxic analogue is also present [46].…”

Caracterización de la fosfatasa PP4 en respuesta a una lesión en el ADN

“…Para las depleciones génicas se amplificaron mediante PCR genes de resistencia a higromicina B (HPH), nourseotricina sulfato (cloNAT) o geneticina (G418) utilizando plásmidos pAG (Euroscarf) [200]. Así mismo, también se utilizaron los genes URA3, TRP1 y LEU2 como marcadores auxotróficos para la depleción de genes en cepas ura3-52 [201], leu2,3-112 [202] o trp1::hisG, respectivamente. Los productos de PCR amplificados contenían secuencias homólogas a las zonas inmediatamente anterior y posterior a la región codificante del gen correspondiente en cada caso, permitiendo de este modo la recombinación y sustitución del gen endógeno por el marcador de resistencia/auxotrófico (Fig.…”